ROLE OF INTRACELLULAR CALCIUM IN GLUCOCORTICOID-EVOKED LYMPHOID CELL APOPTOSIS
description
Transcript of ROLE OF INTRACELLULAR CALCIUM IN GLUCOCORTICOID-EVOKED LYMPHOID CELL APOPTOSIS
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ROLE OF INTRACELLULAR CALCIUM IN
GLUCOCORTICOID-EVOKED
LYMPHOID CELL APOPTOSIS
Devin Morris
California State University, Northridge
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Apoptosis
Release ofcyctochrome c
EffectorCaspases
ApoptosisStimuli/Death signals
Apoptosomeformation
Activation of caspase cascade
Apaf-1 Caspase 9
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Proliferation Cell Population Cell Death
• Development• Homeostasis• Disease
Physiological roles of apoptosis
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R IPC
AB
C
IPCR
IPC
R
RR
RR
ligand receptor
interactingproteins
coactivator complex
basal trans. factors
1
2
3
4
Glucocorticoid Pathway
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Susceptibility of CEM Cell Clones to GC-Evoked Apoptosis
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GC-induced Increase in Intracellular Calcium in GC-Susceptible CEM cells
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Time- & GC-Dose-Dependent Increase in [Ca+2]i in CEM-C7-14 Cells
EtOH 100 nM Dex 1M Dex
24 h
ou
rs48
ho
urs
4.4% 6% 14.1%
5% 24.8% 62.1%
A B C
D E F
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Modulators of [Ca+2]i Levels Influence GC-Evoked Death of CEM Cells
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EGTA Suppresses GC-Evoked Increase in [Ca+2]i Levels in CEM-C7-14 Cells
EtOH 100 nM Dex 1M Dex
Eth
ano
l20
M
EG
TA
A B C
D E F
4.4% 16.4% 35%
2% 12.4% 23.1%
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Glucocorticoid Signaling Pathway
GC [Ca+2]i
(Release fromIntracellular
Stores)
TMB-8
Ca+2-CaM
Binding and Activation
W7 orcalmidazolium
chlorideActivation of
CAMK II
KN-62 orKN-93
Activation of Calcineurin
FK-506 or Cyclosporine A
GC [Ca+2]i
(Release fromIntracellular
Stores)
TMB-8
Ca+2-CaM
Binding and Activation
W7 orcalmidazolium
chlorideActivation of
CAMK II
KN-62 orKN-93
Activation of Calcineurin
FK-506 or Cyclosporine A
Pathway Inhibitors
Normal Pathway
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Inhibition of Calmodulin Protects CEM-C7-14 Cells from GC-evoked Death
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Inhibition of Calmodulin Kinase II Protects CEM-C7-14 Cells from GC-evoked Death
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Inhibition of Calcineurin Protects CEM-C7-14 Cells from GC-evoked Death
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Conclusions
• GCs increase [Ca+2]i levels only in the GC-susceptible CEM-C7-14 cell line in a dose dependent manner; not in the GC-resistant sister cell line, CEM-C1-15.
• Calcium chelation by either BAPTA or EGTA protected CEM-C7-14 cells from GC-evoked apoptosis, in conjunction with a reduction in the amount of free [Ca+2]i.
• The calcium ionophore A23187 causes sensitization of CEM-C1-15 cells to GC-evoked apoptosis.
• Inhibition of calmodulin, calmodulin kinase II or calcineurin, all intermediates in the calcium signaling pathway, impart varying degrees of protection to CEM-C7-14 cells from GC-evoked apoptosis.
• Our data demonstrate a clear correlation between calcium signaling and GC-evoked apoptosis
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Future Goals
• Further studies will aim to understand Ca+2-dependent changes in gene regulation that contribute to apoptosis. Candidate genes such as the transcriptional repressor E4BP4, and its downstream targets are being studied.
• Our ultimate goal is to understand the molecular pathway for apoptosis in T-lymphoid cells as well as in other physiologically relevant models for apoptosis, such as osteoblasts, keratinocytes and macrophages.
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Acknowledgements
Funded by grants from the NIH MBRS-SCORE Program, the CSUN Office of Graduate Studies, Research and International Programs, and the CSUN College of Science & Mathematics.
• Dr. Rheem Medh
• Saul Priceman
• Dr. Carol Shubin
• NASA CSUN/JPL Pair Program